The long-term objectives of this application are to understand the normal function of the Thod gene and to assess its potential suitability as a molecular target for cancer therapy and prognosis. Thod encodes a protein (pThocI) that has recently been identified as a component of the evolutionarily conserved TREX complex. TREX is required for the efficient transcriptional elongation of a subset of genes, and it physically couples elongation to the processes of RNA processing and nuclear export. The yeast orthologue of Thod is not essential for viability, but loss of this gene causes reduced cellular lifespan, reduced growth rate, and increased sensitivity to DNA damage. Similarly, human cancer cells depleted of pThod exhibit reduced growth, viability, and resistance to genotoxic chemotherapeutics. In contrast, normal differentiated cells are relatively unaffected by loss of pThod. The physiological requirements for Thod in normal development and cancer have yet to be assessed in vivo in a multicellular organism. We hypothesize that cancer cells are uniquely dependent on Thod expression for growth and viability, particularly in the presence of DNA damage. The general experimental approach proposed for testing the hypothesis is to measure the effects of pThod loss on normal development and malignant transformation using genetically engineered Thod alleles and autochthonous mouse models of breast cancer. Four specific aims are proposed: 1) Test whether pThod depletion inhibits malignant transformation in vitro. 2) Determine if pThod depletion affects normal mammary gland development. 3) Ascertain whether pThod depletion inhibits breast carcinogenesis in vivo. 4) Assess whether pThod levels influence the response of breast cancer to genotoxic therapy. Cancer cells accumulate genetic and epigenetic alterations that endow them with unwanted proliferative potential, but also burden them with unique vulnerabilities. Thus it is possible to identify genes that are required for. the viability of cancer cells, but not normal cells. Such synthetic lethal genetic interactions identify potential molecular targets for therapy that promise to yield greater specificity for cancer cells. Successful completion of the proposed study will provide proof of principle that Thod is such a molecular target. Since pThod is a component of the newly discovered TREX complex, it functions with a novel mechanism of action. Therapies and diagnostics based on targeting pThod are expected to yield novel clinical responses, opportunities for novel combination therapies, and prognostic information potentially independent of currently used criteria. The proposed research is highly relevant to human health because new molecularly targeted therapies with greater specificity for cancer cells are clearly needed. The experiments proposed will determine whether pThod is a promising target for the development of new therapies and prognostic tests.